Trade‐offs between morphology and thermal niches mediate adaptation in response to competing selective pressures

The effects of climate change—such as increased temperature variability and novel predators—rarely happen in isolation, but it is unclear how organisms cope with multiple stressors simultaneously. To explore this, we grew replicate Paramecium caudatum populations in either constant or variable temperatures and exposed half to predation. We then fit thermal performance curves (TPCs) of intrinsic growth rate (r_max) for each replicate population (N = 12) across seven temperatures (10°C–38°C). TPCs of P. caudatum exposed to both temperature variability and predation responded only to one or the other (but not both), resulting in unpredictable outcomes. These changes in TPCs were accompanied by changes in cell morphology. Although cell volume was conserved across treatments, cells became narrower in response to temperature variability and rounder in response to predation. Our findings suggest that predation and temperature variability produce conflicting pressures on both thermal performance and cell morphology. Lastly, we found a strong correlation between changes in cell morphology and TPC parameters in response to predation, suggesting that responses to opposing selective pressures could be constrained by trade‐offs. Our results shed new light on how environmental and ecological pressures interact to elicit changes in characteristics at both the individual and population levels. We further suggest that morphological responses to interactive environmental forces may modulate population‐level responses, making prediction of long‐term responses to environmental change challenging.     

Analysis of spontaneous and bleomycin-induced chromosome damage in peripheral lymphocytes of long-haul aircrew members from Argentina

Spontaneous and bleomycin (BLM)-induced chromosomal aberrations in G0 and G2 stages of the cell cycle have been analyzed in peripheral lymphocytes of 21 long-haul aircrew members from Argentina in order to assess BLM-induced clastogenesis as a first approach to determine the DNA repair capacity and thereby the susceptibility to environmental cancers in aircrew. The possibility that occupational exposure of flight personnel to cosmic radiation can induce an adaptive response in their peripheral lymphocytes that can be detected by a subsequent in vitro treatment with BLM was also investigated. For comparison, aberrations were also scored in the lymphocytes of 15 healthy volunteers matched by age, health, sex, drinking and smoking habits to the flight personnel group. Aircrew exhibited a higher frequency of spontaneous dicentrics and ring chromosomes than the control population (p<0.05). BLM sensitivity test showed that aircrew and controls are equally sensitive to BLM G2 clastogenic effects, since both groups exhibited a similar frequency of chromatid breaks per cell (p>0.05). However, the aircrew sampled population was almost two times more sensitive to BLM G0 clastogenic effects than controls (p<0.05). Therefore, our data suggest that chronic exposure of aircrew to cosmic radiation increases the in vitro chromosomal sensitivity of their peripheral lymphocytes to BLM (at least in the G0 stage of the cell cycle), and that occupational exposure of flight personnel to cosmic radiation does not induce an adaptive response to this radiomimetic compound. Our results justify further studies aimed at determine if those aircrew members hypersensitive to BLM are more prone to develop environmental cancer than BLM-insensitive individuals.     

Bleomycin Exerts Ambivalent Antitumor Immune Effect by Triggering Both Immunogenic Cell Death and Proliferation of Regulatory T Cells

Bleomycin (BLM) is an anticancer drug currently used for the treatment of testis cancer and Hodgkin lymphoma. This drug triggers cancer cell death via its capacity to generate radical oxygen species (ROS). However, the putative contribution of anticancer immune responses to the efficacy of BLM has not been evaluated. We make here the observation that BLM induces immunogenic cell death. In particular, BLM is able to induce ROS-mediated reticulum stress and autophagy, which result in the surface exposure of chaperones, including calreticulin and ERp57, and liberation of HMBG1 and ATP. BLM induces anti-tumor immunity which relies on calreticulin, CD8⁺ T cells and interferon-γ. We also find that, in addition to its capacity to trigger immunogenic cell death, BLM induces expansion of Foxp3+ regulatory T (Treg) cells via its capacity to induce transforming growth factor beta (TGFβ) secretion by tumor cells. Accordingly, Treg cells or TGFβ depletion dramatically potentiates the antitumor effect of BLM. We conclude that BLM induces both anti-tumor CD8⁺ T cell response and a counteracting Treg proliferation. In the future, TGFβ or Treg inhibition during BLM treatment could greatly enhance BLM anti-tumor efficacy.     

Influence of p53 expression on sensitivity of cancer cells to bleomycin

In this study, we determined whether p53 expression affected the sensitivity of non–small cell lung cancer (NSCLC) and colon cancer cells to bleomycin (BLM). Two human NSCLC cell lines (A549 expressing wild‐type p53 and p53‐null H1299) and two colon cancer cell lines (HCT116 p53+/+ and its p53 deficient subline HCT116 p53?/?) were subjected to treatment with BLM. Cells were treated with various concentrations of BLM, and cellular viability was assessed by formazan assay. To investigate the role of p53 in BLM sensitivity, we transduced cells with adenovirus with wild‐type p53, dominant‐negative p53, and GFP control, and analyzed the effect on cellular viability. Cells expressing wild‐type p53 were more sensitive to BLM than p53‐null cells in both NSCLC and colon cancer cells. Sensitivity to BLM of the cells with wild‐type p53 was reduced by overexpression of dominant‐negative p53, while BLM sensitivity of p53‐null cells was increased by wild‐type p53 in both NSCLC cells and colon cancer cells. In conclusion, our data show that p53 sensitizes all four cancer cells lines tested to BLM toxicity and overexpression of p53 confers sensitivity to the cytotoxic activity of the anticancer agent in p53‐null cells. © 2010 Wiley Periodicals, Inc. J Biochem Mol Toxicol 24:260–269, 2010; View this article online at wileyonlinelibrary.com . DOI 10.1002/jbt.20334     

Testes Investment along a Vertical Depth Gradient in an Herbivorous Fish

Males are expected to adjust testes investment according to the varying level of sperm competition that they experience. Spatial and temporal variation in population density likely influences sperm competition. In herbivorous aquatic organisms, densities often decrease along a vertical depth gradient, because their food is photosynthetic and thus becomes less abundant in deeper regions where less light penetrates. This decrease should be dramatic on a steep slope, which allows testing of the association between density and testes investment at the within‐population level. We tested the effect in the socially monogamous herbivorous cichlid fish Variabilichromis moorii living on a steep slope in Lake Tanganyika. We examined competitor density and food abundance as ecological factors, territory defense behaviors and phenotypic traits (testes investment and somatic investment), and compared them between shallow (4–6 m depth) and deep habitats (10–13 m depth) separated by several dozen meters. We found that food availability drastically decreased with increasing depth and that V . moorii was much more abundant in shallower habitats. Males in shallower habitats were in better physical condition (based on fat and liver mass) despite experiencing greater costs in terms of territory defense. Testes investment differed in areas with different competitor density and food abundance along a vertical depth gradient, but competitor density was the most explanatory factor of the difference. This suggests that this herbivorous fish would change testes investment in response to population density.     

Heat stress adaptation of Escherichia coli under dynamic conditions: effect of inoculum size*

Aims: When subjected to dynamic temperatures surpassing the expected maximum growth temperature, Escherichia coli K12 MG1655 shows disturbed growth curves. These irregular population dynamics were explained by considering two subpopulations, i.e. a thermoresistant and a thermosensitive one ( Van Derlinden et al. 2010a ). In this paper, the influence of the initial cell concentration on the subpopulations’ dynamics is evaluated. Methods and Results: Experiments were performed in a bioreactor with the temperature increasing from 42 to 65·2°C (1 and 4°C h^?1) with varying initial cell concentrations [6, 12 and 18 ln(CFU ml^?1)]. When started from the highest cell concentration, the population was characterized by a higher overall maximum growth temperature and a higher inactivation temperature. For all experimental set‐ups, resistant cells were still growing at the final temperature of 65·2°C. Conclusions: The initial cell concentration had no effect on temperature resistance. The increase in temperature resistance of the sensitive subpopulation was because of the change of the physiological state to the stationary phase. Significance and Impact of the Study: A higher initial cell concentration leads to higher heat stress adaptation when cultures reach a maximum cell concentration. The observed growth at a temperature of 65·2°C is very important for food safety and the temperature treatment of micro‐organisms.     

DNA polymerase III-dependent repair synthesis in response to bleomycin in toluene-treated Escherichia coli

Summary Bleomycin (BLM) is an antitumor drug which interacts with and damages DNA. We have reported a repair response dependent on DNA polymerase I in toluene-treated Escherichia coli. We report here that DNA polymerase III can also catalyze a repair response in toluene-treated E. coli following exposure to BLM. Polymerase III-mediated synthesis differs because it is ATP-dependent, whereas polymerase I-mediated repair synthesis is not. Polymerase III repair synthesis is independent of replicative synthesis, as demonstrated in a polA ^-, dnaB _ts strain, or use of Novobiocin to inhibit replication, and replication persists in the presence of repair synthesis. It appears that ATP-dependent repair synthesis in response to BLM is also present in polA ⁺ strains. Repair synthesis does not require the uvrA gene product.This research was supported by Public Health Service grants GM-19122 and GM-24711 from the National Institute of General Medical Sciences, Robert A. Welch Foundation grant Q-543 and American Cancer Society BC-290     

微地形对大西沟新疆野杏萌发层土壤因子的影响

野杏是新疆野果林的主要建群种和种质资源树种。野杏在种子萌发成苗期,幼苗的根系主要分布在0—15 cm土层中。为了阐明坡向、坡位、坡度和坡形等微地形因子对野杏种子萌发层土壤特征和养分分布的影响,在新疆伊犁州霍城县大西沟的封育野果林内(44°26′01.09″—44°26′17.12″N,80°46′27.49″—80°47′03.26″E)设置样地,测定0—15 cm土层的土壤砾石、酸碱值、有机质和全氮、全磷、全钾、水解性氮、有效磷和速效钾含量,分析各土壤因子和地形因子的关系。结果表明:(1)东北坡土壤养分含量较高,其土壤砾石含量、酸碱值、有机质、全磷、水解性氮和速效钾含量与东南坡、南坡皆存在显著差异(P0.05),阴坡的土壤有机质、全氮、有效磷和速效钾含量都高于阳坡,不同坡向的土壤特征和养分分布存在显著差异(P0.05);(2)不同坡位的土壤有机质、全氮、全磷和有效磷含量均表现为中坡位下坡位上坡位,土壤砾石、全钾和速效钾含量均呈现上坡位中坡位下坡位,全钾、全磷和有效磷含量在同坡位分布较均匀,不同坡位的土壤特征和土壤养分分布无显著差异(P0.05);(3)不同坡度下的土壤水解性氮和有效磷含量是缓坡中坡缓中坡陡坡,土壤砾石、酸碱值、全氮、水解性氮、有效磷和速效钾含量有显著差异(P0.05),陡坡土壤有机质、全氮、水解性氮和有效磷含量均低于其他坡度,而土壤砾石、土壤酸碱值和全钾在陡坡土层中含量最高,缓坡与缓中坡的土壤养分含量丰富,陡坡较为贫瘠,坡度对土壤特征和土壤养分分布有显著影响(P0.05);(4)不同坡形下的土壤砾石、全钾、水解性氮、有效磷和速效钾含量是凸形坡凹形坡直线坡,不同坡形下的土壤特征和土壤养分(除全钾和速效钾)均存在显著差异性(P0.05)。野果林地形因子对野杏萌发层土壤特征和土壤养分分布有显著影响,东北坡、中坡位、缓坡和凸形坡土壤养分含量较为丰富,是适宜野杏萌发的地形。研究结果可为探究影响野杏种子萌发和生长的气候与土壤水热等因子奠定良好基础。     

微地形对江西武夷山南方铁杉针阔混交林幼树更新的影响

为探讨江西武夷山南方铁杉针阔混交林内不同微地形中幼树更新特征的差异,明晰不同微地形生境对幼树天然更新的影响。以江西武夷山国家级自然保护区内海拔约1800m的南方铁杉针阔混交原始林为研究对象,基于在其中建立的中亚热带南方铁杉针阔混交林动态监测固定样地首次调查数据,将固定样地中160个20m×20m的样方根据其海拔、凹凸度和坡度3种地形参数通过C-均值模糊聚类划分成不同的微地形生境,比较这不同微地形下的更新幼树种类组成及幼树更新特征,包括幼树密度,幼树平均胸径、平均高、平均冠幅,空间分布格局,分析不同微地形下幼树更新特征与地形因子、林分因子之间的关系。结果显示：(1)通过聚类分析,最终将固定样地的微地形生境划分成4类,分别为陡坡、凹地、凸地、缓坡。(2)4类微地形更新幼树优势种组成类似,但优势程度存在差异。闽皖八角幼树在4类微地形中的重要值皆为第一,均大于43%。建群种南方铁杉的幼树在凸地的重要值为9.09%,具有一定优势;在陡坡的重要值为0.62%,优势程度较差。(3)陡坡的更新幼树密度最高,为399株/hm~2,高于凹地,凸地和缓坡;幼树平均胸径和平均高的大小表现为：缓坡>凸地&...     

Biomass partitioning in twoCalamagrostis villosa stands on deforested sites

Calamagrostis villosa stands occurring in areas deforested by air-pollution impact in the Moravian-Silesian Beskydy Mountains were characterized by a high dry mass of total underground biomass (3 300 g. m^?2—the slope site, 2 850 g. m^?2—the flat site). The percentage of living roots and rhizomes in total underground biomass was very high (about 70%). The total aboveground biomass was respectively, 321 g.m^?2 (the slope site) and 726 g. m^?2 (the flat site). In unstabilized habitats on steep slope, the higher plant biomass produced was allocated to a more developed root system.     

Epithelial vs myofibroblast differentiation in immortal rat lung cell lines—modulating effects of bleomycin

Two alveolar epithelial cell lines R3/1 and L2 were screened by immunocytochemical and RT-PCR analysis of epithelial and mesenchymal/contractile marker proteins. R3/1 and L2 cells were tested for their sensitivity to bleomycin (BLM), an anticancer drug, which is proposed to induce changes in lung cell differentiation. Both epithelial cell lines exhibited a mixed phenotype consisting of epithelial (E-cadherin, aquaporin-5 and cytokeratin 8) and myofibroblast-like (vimentin, α-SMA and caveolin-3) properties suggesting that the cell lines are arrested in vitro at a certain developmental stage during epithelial–mesenchymal transition (EMT). BLM treatment of R3/1 cells resulted in a partial reversal of this process modifying the cells in an epithelial direction, e.g., upregulation of E-cadherin, aquaporin-5 and other lung epithelial antigens at the mRNA and protein level. L2 cells showed similar alterations following BLM exposure. Immunohistochemical investigation of lung tissue from two different animal models of BLM-induced fibrosis (mouse and rat), revealed no signs of EMT, e.g., myofibroblastic differentiation of alveolar epithelial cells in situ. Immunohistological analysis of tissue samples of the rat model showed a heterogeneous population of myofibroblasts (α-SMA⁺/caveolin-3⁺, α-SMA^-/caveolin-3⁺, and α-SMA⁺/caveolin-3⁻). These results suggest that BLM, on one hand, induces fibrosis and on the other hand possibly suppresses EMT during fibrogenesis.     

Repair response of human fibroblasts to bleomycin damage

The ability of human fibroblasts to repair the specific types of DNA damage caused by bleomycin (BLM) was examined in whole-cell experiments. The method utilized for analysis was alkaline sucrose-gradient centrifugation of DNA. The results of these studies show that a repair pathway exists for the damage produced in DNA by bleomycin. DNA from BLM-treated cells shows a decrease in molecular weight, caused by chemical or enzymatic incision at sites of drug action. If the drug is removed, the DNA rapidly returns to high molecular weight, demonstrating reformation of damaged DNA. This repair response to BLM-damage was also confirmed in fibroblasts isolated from patients with putative DNA-repair defects. We observed that the response (to BLM) of cells from patients with Fanconi anemia was altered in that the fall in molecular weight of DNA from treated cells was not as great as that observed in other cell strains after drug treatment.     

Up-regulation of antioxidant enzymes and coenzyme Q10 in a human oral cancer cell line with acquired bleomycin resistance

Abstract

Bleomycin (BLM) is an anti-cancer drug that can induce formation of reactive oxygen species (ROS). To investigate the association between up-regulation of antioxidant enzymes and coenzyme Q₁₀ (CoQ₁₀) in acquired BLM resistance, one BLM-resistant clone, SBLM24 clone, was selected from a human oral cancer cell line, SCC61 clone. The BLM resistance of SBLM24 clone relative to a sub-clone of SCC61b cells was confirmed by analysis of clonogenic ability and cell cycle arrest. CoQ₁₀ levels and levels of Mn superoxide dismutase, glutathione peroxidase 1, catalase and thioredoxin reductase 1 were augmented in SBLM24 clone although there was also a mild increase in the expression of BLM hydrolase. Suppression of CoQ₁₀ levels by 4-aminobenzoate sensitized BLM-induced cytotoxicity. The results of suppression on enhanced ROS production by BLM and the cross-resistance to hydrogen peroxide in SBLM24 clone further demonstrated the development of adaptation to oxidative stress during the formation of acquired BLM resistance.     

N-acetylcysteine amide,a thiol antioxidant,prevents bleomycin-induced toxicity in human alveolar basal epithelial cells (A549)

Abstract

Bleomycin (BLM), a glycopeptide antibiotic from Streptomyces verticillus, is an effective antineoplastic drug. However, its clinical use is restricted due to the wide range of associated toxicities, especially pulmonary toxicity. Oxidative stress has been implicated as an important factor in the development of BLM-induced pulmonary toxicity. Previous studies have indicated disruption of thiol-redox status in lungs (lung epithelial cells) upon BLM treatment. Therefore, this study focused on (1) investigating the oxidative effects of BLM on lung epithelial cells (A549) and (2) elucidating whether a well-known thiol antioxidant, N-acetylcysteine amide (NACA), provides any protection against BLM-induced toxicity. Oxidative stress parameters, such as glutathione (GSH), malondialdehyde (MDA), and antioxidant enzyme activities were altered upon BLM treatment. Loss of mitochondrial membrane potential (ΔΨm), as assessed by fluorescence microscopy, indicated that cytotoxicity is possibly mediated through mitochondrial dysfunction. Pretreatment with NACA reversed the oxidative effects of BLM. NACA decreased the reactive oxygen species (ROS) and MDA levels and restored the intracellular GSH levels. Our data showed that BLM induced A549 cell death by a mechanism involving oxidative stress and mitochondrial dysfunction. NACA had a protective role against BLM-induced toxicity by inhibiting lipid peroxidation, scavenging ROS, and preserving intracellular GSH and ΔΨm. NACA can potentially be developed into a promising adjunctive therapeutic option for patients undergoing chemotherapy with BLM.     

Orientation of the sand-beach amphipod, Orchestoidea corniculata

Laboratory experiments demonstrated that Orchestoidea corniculata, a sand-beach amphipod, can orient to slopes of only 3°. Studies suggest they reach the upper interdial zone where they burrow by moving up a wet slope or down a dry slope. When they emerge from the sand at night, however, they either reverse this response or show no response to slope. This behaviour, enhanced by a preference for wet, surface sand, would account for the initial seaward movement observed in the field. Behavioural variations among differing segments of the population were investigated. Three additional beach inhabitants oriented to slope, thus indicating the mechanism of slope orientation as a possible generalization in the beach habitat.     

Modulation of bleomycin-induced mitotic recombination in yeast by the aminothiols cysteamine and WR-1065

The cancer chemotherapy drug bleomycin (BLM) is a potent inducer of genetic damage in a wide variety of assays. The radioprotectors cysteamine (CSM) and WR-1065 have been shown in previous studies to potentiate the induction of micronuclei and chromosome aberrations by BLM in Go human lymphocytes. By contrast, WR-1065 is reported to reduce the induction of hprt mutations by BLM in Chinese hamster cells. To elucidate the basis for these interactions, we examined the effects of CSM and WR-1065 on the induction of mitotic gene conversion by BLM in the yeast Saccharomyces cerevisiae. Treatment with BLM causes a dose-dependent increase in the frequency of mitotic gene conversion and gene mutations. Unlike its potentiation of BLM in G₀ lymphocytes, WR-1065 protected against the recombinagenicity of BLM in yeast. CSM was also strongly antirecombinagenic under some conditions., but the nature of the interaction depended strongly on the treatment conditions. Under hypoxic conditions, cysteamine protected against BLM, but under oxygenrich conditions CSM potentiated the genetic activity og BLM. The protective effect of aminothiols against BLM may be ascribed to the depletion of oxygen required for the activation of BLM and the processing of BLM-induced damage. Aminothiols may potentiatc the effect of BLM by acting as an electron source for the activation of BLM and/or by causing conformational alterations that make DNA more accessible tc BLM. The results indicate that aminothiols have a strong modulating influence on the genotoxicity of BLM in yeast as they do in other genetic assays. Moreover, the modulation differs markedly depending on physiological conditions. Thus, yeast assays help to explain why aminothiols have been observed to potentiate BLM in some genetic systems and to protect against it in others.     

BLM and the FANC proteins collaborate in a common pathway in response to stalled replication forks

Fanconi anaemia (FA) and Bloom syndrome (BS) are autosomal recessive diseases characterised by chromosome fragility and cancer proneness. Here, we report that BLM and the FA pathway are activated in response to both crosslinked DNA and replication fork stall. We provide evidence that BLM and FANCD2 colocalise and co-immunoprecipitate following treatment with either DNA crosslinkers or agents inducing replication arrest. We also find that the FA core complex is necessary for BLM phosphorylation and assembly in nuclear foci in response to crosslinked DNA. Moreover, we show that knock-down of the MRE11 complex, whose function is also under the control of the FA core complex, enhances cellular and chromosomal sensitivity to DNA interstrand crosslinks in BS cells. These findings suggest the existence of a functional link between BLM and the FA pathway and that BLM and the MRE11 complex are in two separated branches of a pathway resulting in S-phase checkpoint activation, chromosome integrity and cell survival in response to crosslinked DNA.     

Environmental adaptation and genetic variations in geographically isolated Emma field crickets Teleogryllus emma (Orthoptera: Gryllidae)

The Emma field cricket, Teleogryllus emma (Ohmachi & Matsuura), distributed between 43°N and 30°N in the Japanese archipelago, is univoltine and overwinters in the egg stage. Its eggs hatch on the slope of the Oishi Dam (38.03°N, 139.57°E, 160–170 m a.s.l.) in late June, adults begin emerging from late August, and oviposition lasts until early October. Oviposition is limited to the period when the water level of the Oishi Dam is low. The period from egg hatching to adult emergence is approximately 1 month shorter than that of the T. emma population on the Arakawa riverside (38.09°N, 139.57°E, 29 m a.s.l.), which is approximately only 7 km from the Oishi Dam. The egg and body sizes of T. emma on the slope of the Oishi Dam were smaller than those of T. emma on the Arakawa riverside, and the egg and nymphal periods were shorter; these variations were inherited by the next generation of T. emma. The egg period, nymphal period and head width of T. emma on the dam slope correspond to those of the populations near 40°N. Several traits of the T. emma population on the dam slope were naturally selected by adapting to the isolated environment, resulting in the genetic variations. However, their variations were small and the period after isolation is short, suggesting that it is in the early stages of speciation.     

USP37 regulates DNA damage response through stabilizing and deubiquitinating BLM

The human RecQ helicase BLM is involved in the DNA damage response, DNA metabolism, and genetic stability. Loss of function mutations in BLM cause the genetic instability/cancer predisposition syndrome Bloom syndrome. However, the molecular mechanism underlying the regulation of BLM in cancers remains largely elusive. Here, we demonstrate that the deubiquitinating enzyme USP37 interacts with BLM and that USP37 deubiquitinates and stabilizes BLM, thereby sustaining the DNA damage response (DDR). Mechanistically, DNA double-strand breaks (DSB) promotes ATM phosphorylation of USP37 and enhances the binding between USP37 and BLM. Moreover, knockdown of USP37 increases BLM polyubiquitination, accelerates its proteolysis, and impairs its function in DNA damage response. This leads to enhanced DNA damage and sensitizes breast cancer cells to DNA-damaging agents in both cell culture and in vivo mouse models. Collectively, our results establish a novel molecular mechanism for the USP37–BLM axis in regulating DSB repair with an important role in chemotherapy and radiotherapy response in human cancers.     

Redox potential of the active bleomycin-Fe(III) complex by cyclic voltammetry

The redox potential of the active Fe(III) complex of bleomycin (BLM), which is a DNA cleaving species, was measured by cyclic voltammetry at 25 °C under a hydrogen atmosphere. The cyclic voltammogram showed the reversible one-electron Fe(III)/Fe(II) coupled redox reaction at −0.225 V versus SCE. Under the same conditions the redox potentials of the iso-BLM—Fe(III) complex and the deglyco-BLM—Fe(III) complex were also observed, but the cyclic voltammogram for the inactive Fe(III) complex of BLM could not be obtained.